Volume overload (VO) in the heart promotes bioenergetic, structural, and functional changes that lead to heart failure. However, the mechanisms of myocardial decompensation and the resulting progression to failure due to VO remain unclear. It is established that VO increases xanthine oxidase (XO) and reactive oxygen and nitrogen species (ROS/RNS). In this proposal we will integrate two critical basic research findings in the context of cardiac failure due to VO: 1) XO as a source of oxidative damage in the heart due to VO and 2) the central role mitochondrial dysfunction in the etiology of VO-mediated heart failure. This concept will be tested through pursuit of the following specific aims in targeted animal and human studies. Aim 1 will test the hypothesis that increased XO activity in VO causes LV dysfunction through cardiomyocyte mitochondrial dysfunction in chronic VO in rat. Aim 2 will test the hypothesis that cardiomyocyte-derived XO activity is responsible for cardiomyocyte MMP activation and mitochondrial dysfunction using gene therapy to knockdown XDH in acute and chronic VO in rats. Aim 3 will test the hypothesis that XO inhibition reduces cardiomyocyte oxidative stress and mitochondrial dysfunction and improves LV function after closure of chronic ACF in rat. These experiments will utilize novel gene therapy techniques in rats to prove cause and effect of XO in cardiomyocytes. Studies of cardiomyocyte mitochondrial function will be performed using Seahorse XF24 in animal cells. The animal and cardiomyocyte studies proposed will determine whether increased XO is a key regulator of oxidative stress and mitochondrial dysfunction in the chronic stretch of a pure VO. These studies, if positive, could provide the scientific impetus for a clinical trial of XO inhibition in patients with isolated VO. PUBLIC HEALTH RELEVANCE: Mechanisms of left ventricular dysfunction in models of volume overload are not well understood and there is currently no recommended medical therapy. Using targeted studies in a clinically relevant animal model the principal investigator addresses the novel hypothesis that cardiomyocyte-derived xanthine oxidase mediates oxidative damage of mitochondria and left ventricular dysfunction in volume overload. These studies in animals, if positive, could provide the scientific impetus for a clinical trial of xanthine oxidase inhibition in patients with isolated volume overload.